Repair of acute damage to the plasma membrane is an important aspect of normal cellular physiology and disruption of this process can result in pathophysiology in a number of human diseases including muscular dystrophy. We recently discovered that MG53, a muscle-specific TRIM-family protein, is an essential component of the membrane repair machinery. While our published data define the sensor function for MG53 in cell membrane repair, the molecular mechanisms underlying the nucleation process remain to be defined. Since MG53 can discriminate between intact and injured membrane, a membrane-delimited signal would likely be involved in tethering of MG53 to the injured site. In pilot studies, we found that PTRF is an obligatory factor for MG53-mediated nucleation of the membrane repair response, for cells lacking endogenous expression of PTRF show defective membrane resealing. While RNAi- silencing of PTRF leads to defective membrane repair in muscle fibers, overexpression of PTRF can rescue this defect in dysferlin-/- muscle but not in mg53-/- muscle, suggesting that the functional role of PTRF in membrane repair likely requires the presence of MG53. While many studies have explored the function of PTRF in regulating caveolae structure of the plasma membrane, our data present a new biological function for PTRF as an anchoring molecule for MG53 for initiation of the cell membrane repair response. Since mutations in PTRF have been identified in human disorders with lipodystrophy and muscular dystrophy, conditions that often involve compromised membrane integrity or resealing capacity, targeting the functional interaction between MG53 and PTRF, or restoration of the disrupted MG53-PTRF interaction in the diseased states, may represent an attractive avenue for treatment or prevention of degenerative diseases involving compromised membrane repair. The long-term goal of this project is to understand the cellular and molecular mechanism for membrane repair in muscle physiology and diseases. Specifically, we will focus on testing the hypothesis that PTRF acts as a docking protein for MG53-mediated cell membrane repair, and restoration of membrane integrity in muscular dystrophy can be achieved through enhancement of MG53/PTRF function at the interface of membrane injury. Our proposed studies will focus on defining the molecular mechanism underlying the functional interaction between MG53 and PTRF for initiation of the cell membrane repair response in skeletal muscle (Aim 1); and exploring the physiological role of MG53 and PTRF in muscle physiology and diseases and test if enhancement of PTRF-MG53 function can improve membrane integrity in muscular dystrophy (Aim 2). Through tailored-expression of MG53 and PTRF and the use of biochemical markers, live cell imaging, ex vivo and in vivo animal model studies, the designed experiments will provide key proof-of-principle data for targeting MG53/PTRF-mediated cell membrane repair in treatment of muscular dystrophy.